Analysis of connexin26 role in mouse skin carcinogenesis.. The primary objective of this project is to study a fundamental and novel molecular mechanism of cell growth control by gap junctional proteins, the connexins (Cx). Considerable evidence indicates that connexin genes are tumor suppressor genes since loss or change of connexin function provides a growth advantage to cells. However, it is well understood now that some connexins, especially connexins from so-called alpha and beta groups are incompatible. Hemi-channels composed from incompatible connexins do not form functional channels. This led us to the hypothesis that Co- expression of incompatible connexins may actually decrease gap junctional intercellular communication (GJIC), thereby releasing cells from connexin-mediated growth control during carcinogenesis. The primary objective of this project is to test this innovative hypothesis using as a model co-expression of incompatible Cx26 (beta2) and Cx43 (alpha1) in keratinocytes. The comprehensive analysis of connexin expression in normal mouse skin and in skin tumors, by my laboratory, indicated an exceptional role for Cx26/Cx43 combination during skin tumor promotion. Keratinocytes in adult mouse epidermis express several connexins including Cx43 but not Cx26. In 90% of skin papillomas we observed the overall simplification of connexin expression profile with a shift towards expression of only two major connexins, Cx26 and Cx43, which were co- localized on the plasma membranes of transformed keratinocytes. To investigate the causative role of Cx26/Cx43 Co-expression in the pathogenesis of skin tumors, we designed the following Specific Aims: 1) Determine whether co-expression of incompatible connexins Cx26 and Cx43 inhibits GJIC in keratinocytes; 2) Determine whether co-expression of incompatible connexins Cx26 and Cx43 provides a growth advantage to keratinocytes; 3) Develop transgenic animals deficient in Cx26 expression in epidermis, and Cx26 transgenics with Cx26 targeted to Cx43-positive basal keratinocytes. The results of these experiments will have fundamental importance in deciphering the alternative mechanisms of growth control by connexins.